This project deals with the analysis of regulation of eucaryotic nuclear genes. The regulation of nuclear genes is instrumental in orchestrating the various developmental patterns which occur during differentiation and the defects of which are reflected in diseases such as cancer. For the analysis of eucaryotic nuclear genes we are utilizing a set of yeast genes which code for the biosynthetic enzymes involved in the synthesis of isoleucine, valine, and leucine. This gene system called ilv is composed of five different structural genes which are not linked on the same chromosome and which are regulated by various end products of the pathways, namely, isoleucine, valine, and leucine. Genetic analysis of the ilv 1 gene indicates that there may be two proteins encoded by the gene. One protein specific for the biosynthetic activity threonine deaminase and the other protein appears to be involved in the regulation of the ilv 2, 3, and 4 genes. This project involves the analysis of this gene system and the development of various new techniques to take advantage of the different biochemical and genetic properties of the ilv system. We have improved the yeast transformation procedure which is instrumental for isolating the various ilv genes. The availability of purified genes, of course, permit their analysis by physical techniques. In addition, we are developing a technique which permits the isolation of specific DNA binding proteins such as proteins involved in the regulation of the various ilv genes.